The objective of this research is to learn more about the way in which the development of nerve cells proceeds. The sequence in which some different cellular phenotypes are expressed, as a presumptive nerve cell becomes a fully differentiated neuron, has been determined for amphibian spinal neurons growing in vivo and in vitro. The birthdate and ultrastructural development of two of the three neuronal types of earliest origin, the Rohon-Beard neurons, the extramedullary neurons and the primary motor neurons have been studied. The cells extend neurites at very early stages and the development of three different membrane properties has been examined (electrical excitability, chemosensitivity, and high affinity neurotransmitter uptake). These cells will be further studied in culture in this analysis of neuronal development. The project is designed to answer some questions about the timing of the RNA and protein syntheses involved in the acquisition of each of these phenotypes. RNA and protein synthesis will be inhibited by continuous application of specific drugs, starting at later and later stages of development, and the time of drug application at which RNA and protein synthesis inhibitors no longer prevent the expression of electrical excitability, chemosensitivity, and high affinity neurotransmitter uptake will be determined. These experiments will provide the relation between the time of RNA synthesis, protein synthesis, and expression for each phenotype studied. Further, this project will answer questions about the relationship of the events involved in the expression of one phenotype, to the events involved in the expression of the others. If the periods of RNA or protein synthesis for the neuronal characteristics studied are sufficiently separate, then brief application and washout of inhibitors of RNA and protein synthesis will permit experiments in which only a single phenotype is blocked, and allow examination of the consequences for the expression of the others. These studies should permit distinction between simultaneous and sequential syntheses of RNA and protein underlying the expression of these neuronal phenotypes during development.